U2AF65 adapts to diverse pre-mRNA splice sites through conformational selection of specific and promiscuous RNA recognition motifs
Document Type
Journal ArticlePublication Date
2013-04-01Keywords
Nuclear ProteinsRibonucleoproteins
Nucleic Acid Conformation
RNA Splice Sites
Biochemistry, Biophysics, and Structural Biology
Genetics and Genomics
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Degenerate splice site sequences mark the intron boundaries of pre-mRNA transcripts in multicellular eukaryotes. The essential pre-mRNA splicing factor U2AF(65) is faced with the paradoxical tasks of accurately targeting polypyrimidine (Py) tracts preceding 3' splice sites while adapting to both cytidine and uridine nucleotides with nearly equivalent frequencies. To understand how U2AF(65) recognizes degenerate Py tracts, we determined six crystal structures of human U2AF(65) bound to cytidine-containing Py tracts. As deoxy-ribose backbones were required for co-crystallization with these Py tracts, we also determined two baseline structures of U2AF(65) bound to the deoxy-uridine counterparts and compared the original, RNA-bound structure. Local structural changes suggest that the N-terminal RNA recognition motif 1 (RRM1) is more promiscuous for cytosine-containing Py tracts than the C-terminal RRM2. These structural differences between the RRMs were reinforced by the specificities of wild-type and site-directed mutant U2AF(65) for region-dependent cytosine- and uracil-containing RNA sites. Small-angle X-ray scattering analyses further demonstrated that Py tract variations select distinct inter-RRM spacings from a pre-existing ensemble of U2AF(65) conformations. Our results highlight both local and global conformational selection as a means for universal 3' splice site recognition by U2AF(65).Source
Nucleic Acids Res. 2013 Apr 1;41(6):3859-73. doi: 10.1093/nar/gkt046. Link to article on publisher's site
DOI
10.1093/nar/gkt046Permanent Link to this Item
http://hdl.handle.net/20.500.14038/44008PubMed ID
23376934Related Resources
Link to article in PubMedRights
Copyright The Author(s) 2013. Published by Oxford University Press.
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
ae974a485f413a2113503eed53cd6c53
10.1093/nar/gkt046